Pneumocystis carinii causes severe pneumonia in immunocompromised patients with AIDS. Recent investigations have advanced our understanding of host- organism interactions during P. carinii pneumonia. These studies indicate that attachment of P. carinii to alveolar epithelial cells is an integral event which promotes completion of the organism's life cycle. Continuous culture of P. carinii in a cell-free environment continues to be elusive. However, the organism can multiply on cultured respiratory epithelial cells. Attachment of P. carinii trophozoites to epithelial cells occurs by the close apposition of the organism's cell membrane with the membrane of the host. Further studies demonstrate that the attachment of P. carinii to host cells is largely potentiated by the adhesive proteins vitronectin (VN) and fibronectin (FN) present in the alveolar space. However, the role of these attachment mechanisms in promoting completion of the organism's life cycle have not been fully defined. In addition, the attachment of P. carinii to the host, also suppresses division of respiratory epithelial cells, thereby slowing repair of lung injury. The mechanisms by which P. carinii attachment promotes completion of the organism's life cycle, while simultaneously impairing the cell cycle of the host are largely unknown. The eukaryotic cell cycle is carefully regulated by a group of cell division cycle (cdc) molecules including cyclins and cyclin-dependent kinases. Of critical importance is the cdc2 gene which encodes a serine/threonine kinase. The timing of cdc2 activation is controlled in part by the cdc25 gene which activates the cdc2 gene product in late G2. Homologues of cdc2 and cdc25 have been identified in a variety of eukaryotic organisms from Schizosaccharomyces pombe, a fungus closely related to P. carinii, through man. To date, very little has been learned about cell cycle control molecules in P. carinii. Recently, our laboratory has demonstrated that P. carinii possesses a protein containing the PSTAIR amino-acid sequence conserved in cdc2 gene products in all eukaryotes. We hypothesize that attachment of P. carinii to respiratory epithelial cells promotes life cycle completion and proliferation of P. carinii. We propose that P. carinii attachment by VN and FN-mediated mechanisms alters the expression and/or activity of homologous cdc2 and cdc25 gene products in P. carinii. We further postulate that P. carinii attachment also inhibits the cell cycle of host epithelial cells, through altered activity of the mammalian cyclins and cyclin dependent kinases. We will evaluate whether attachment of P. carinii to respiratory epithelial cells through the adhesive proteins VN and FN promotes life cycle progression in the organism. Further, we will identify specific P. carinii cell cycle control molecules using a complementation cloning strategy. Mutants of S. pombe which lack cdc2 and cdc2S have been derived and successfully used to clone homologous molecules from other species. Therefore, we will use complementation to clone the P. carinii homologues of cdc2 and cdc25. Next, the expression and activity of these cell cycle control-molecules will be studied in purified populations of P. carinii trophozoites and cysts. Finally, we will determine the mechanisms by which P. carinii attachment to epithelial cells mediates alterations in the cell cycles of both the organism and the host promises to yield new therapeutic insights in P. carinii pneumonia.